Summary of proposed work: a) Mechanisms of antibiotic resistance in clinical isolates of gram-negative and gram-positive bacteria. New antibiotic resistance mechanisms will be characterized genetically and biochemically; particular attention will be paid to the presence of R factors and the relationships between similar resistance mechanisms in different organisms. b) A study of temperature-sensitive (heat and cold) mutants in S. cerevisiae. A large number of mutants have been isolated; in particular we will study those mutants with a lesion in the protein synthetic apparatus, using a cell-free protein synthesizing system. c) Studies with RNA polymerase inhibitors in S. cerevisiae; the mechanisms of action of several inhibitors including thiolutin, chaetocin, and lomofungin will be studied. Mutants resistant to these inhibitors will be analyzed. d) The methylation of 16S RNA in ribonucleoprotein particles by the ksg methylase will be studied. In particular we wish to determine what ribosomal proteins are required for substrate activity and what ribosomal proteins can inhibit the action of the methylase. e) The mode of action of actinobolin on E. coli, and cryptopleurine on S. cerevisiae will be studied.